Sleep is integral to health, and humans should be asleep for a third of their life. Poor or insufficient sleep raises the risk of a number of pathologies including cardiovascular disease. However, the underlying biological mechanisms that link sleep to cardiovascular health are unclear. Leukocytes of the myeloid lineage, sourced from bone marrow (BM) hematopoiesis, play a central role in cardiovascular pathology, including atherogenesis. Sleep fragmentation activates hematopoietic stem and progenitor cells (HSPCs) resulting in monocytosis and enlarged murine atherosclerotic lesions. The proposed research aims to extend this observation and explore how sleep shapes the epigenome of hematopoietic stem cells, their clonal expansion, lineage commitment, and innate immune entrainment. Dr. Cameron McAlpine has developed innovative mouse models of sleep fragmentation and disruption. Based on published and unpublished data, this proposal hypothesizes that adequate sleep programs the epigenome of leukocytes, promoting an epigenetic profile that fosters HSPC diversity, shapes HSPC lineage commitment, and influences innate immune entrainment. To achieve these aims, innovative mechanical and genetic mouse models of sleep fragmentation will be used. Using a fluorescent HSPC tagging system, the population dynamics of specific HSPC clones will be monitored during sleep fragmentation. Further, HSPC clones that are dynamically regulated by sleep will have their epigenome and transcriptome profiled. Additionally, using single-cell RNA-seq, the influence of sleep on the entire BM HSPC compartment and its cell clusters will be analyzed. Using models of clonal hematopoiesis targeting somatic mutations in epigenetic modifiers, the impact of sleep on clonal hematopoiesis driven myeloproliferation and atherosclerosis will be determined. Finally, how sleep shapes innate immune entrainment caused by diverse inflammatory stimuli will be questioned. Together, this proposed research will provide a comprehensive view on the importance of sleep and its impact on hematopoiesis, the innate immune system, and cardiovascular disease. Dr. Cameron McAlpine will conduct this work within the Center for Systems Biology at the Massachusetts General Hospital and Harvard Medical School under the primary mentorship of Dr. Filip Swirski and co-mentorship of Dr. David Scadden. Dr. McAlpine has assembled an Advisory Committee comprising of Dr. Peter Libby, clinical cardiologist and expert in clonal hematopoiesis; Dr. Kate Jeffery, an expert in leukocyte epigenetics; and Dr. Thomas Scammell, sleep clinician and expert on the neurobiology of sleep. These mentors will allow him to develop a successful research strategy and career as an independent biomedical investigator.